The long-term goal of this project is to understand the parameters that control T cell behavior during development and during immune responses, with a particular focus on the mechanisms that maintain self- tolerance. Previously, we determined that the tumor suppressor gene PTEN, a lipid phosphatase which reverses the PI-3 kinase reaction, is expressed in naTve and regulatory T cells. Using mice with PTEN deletion targeted to the T cell compartment (PTEN-AT mice), we found that PTEN regulates both TCR and IL-2R mediated responses. In the absence of PTEN, naive T cells can be optimally activated in vitro and in vivo without CD28 costimulation, and regulatory T cells can be induced to proliferate to IL-2 without a need for TCR activation. Thus PTEN is a critical regulator of T cell homeostasis. As they age PTEN-AT mice develop T cell lymphomas and a generalized autoimmune syndrome. Previous models suggested that this was due to the lipid phosphatase activity of PTEN, which modulated TCR and growth factor receptor signaling. However new data indicates that PTEN has a nuclear function as well, where it binds to centromeres and mediates chromosomal stability. Interestingly, our preliminary data indicate that lymphoma development is an intrathymic event while autoimmunity arises from mature peripheral T cells, suggesting that distinct mechanisms govern these two consequences of PTEN-deficiency in T cells. Based on these data, we have three aims. In aim #1, we will use define the requirements for malignant transformation and autoimmune potential of PTEN-deficient T cells. This will be done by creating chimeric mice with mutated PTEN constructs that have isolated loss of phosphatase or chromosomal stability domains, and by crossing PTEN-AT mice onto TCR transgenic RAG-deficient, TCRa-deficient, and conditional SLP-76 deficient backgrounds. In aim #2, we will use foxp3-GFP-Cre reporter mice to delete PTEN in the major subset of Tregs, to determine how the loss of PTEN in vivo Treg homeostasis, conversion, and stability, and to test if PTEN-deficient Tregs modulate autoimmune susceptibility. Finally, in aim #3 we will determine how PTEN-deficiency affects the T cell response to a pathogen. Here, we will use Toxoplasma gondii expression ovalbumin as a transgene, and MHC-tetramers (to OVA) as well as OVA- TCR transgenic PTEN-AT mice to follow the fate and migration of labeled PTEN-AT T cells in response to infection. RELEVANCE (See instructions): Cells known as T lymphocytes are critical components of the immune system which are required for successful defense against many types of infections. The research proposed will examine how a gene called PTEN, which is known to be involved in controlling cell growth and cancer, regulates the development and function of T lymphocytes, preventing leukemia and autoimmune disease.